The invention relates generally to a method for modulating intraocular pressure (IOP) in an eye of a human or a non-human mammalian subject having or susceptible to steroid-induced glaucoma, and more particularly to a method for modulating an actin cytoskeleton structures in trabecular meshwork cells in a subject having steroid-induced glaucoma.
The front of a mammalian eye has an anterior chamber, bounded at the front by a cornea and at the back by a lens, as well as a posterior chamber located behind an iris but in front of the lens. Behind the iris is a ciliary body that continuously produces aqueous humor, a thin, watery fluid that fills the anterior and posterior chambers. The aqueous humor nourishes the cornea and the lens and gives the front of the eye its form and shape. The aqueous humor flows from the ciliary body through the pupil and through a trabecular meshwork then through the canal of Schlemm. The trabecular meshwork is the complex tissue located in the angle between the cornea and the iris that contains specialized endothelial cells, connective tissue beams and extracellular matrix and that is responsible for a majority of the resistance to aqueous humor outflow through the anterior chamber angle. In a healthy eye, the aqueous humor under resistance from the structures of the eye, notably the trabecular meshwork, generates normal IOP in a range from about 12-20 mmHg.
When aqueous humor cannot drain normally from the anterior chamber of an eye, an animal can develop one of a family of ophthalmologic disorders characterized by above-normal IOP and gradual neuropathy caused in some manner by increased pressure on the optic nerve. Pressure increase begins in the anterior chamber of the eye and extends to the other parts of the eye, including the posterior chamber. Under the force of the IOP, a posterior segment compresses and destroys nerve fibers and blood vessels of the optic nerve. Such disorders can lead to gradual visual impairment and are collectively referred to as glaucoma.
Of particular interest is steroid-induced glaucoma. In steroid-induced glaucoma, aqueous humor builds up in the anterior chamber because it cannot flow through the trabecular meshwork, thereby increasing IOP. The exact pathophysiology of steroid-induced glaucoma is unknown, but it is known that steroids can increase or decrease transcription of genes, including genes of the trabecular meshwork cells. Ishibashi T., et al., “cDNA microarray analysis of gene expression changes induced by dexamethasone in cultured human trabecular meshwork cells,” Invest. Ophthalmology & Visual Sci. 43:3691-3697 (2002). Other putative mechanisms for steroid-induced glaucoma include the following: (1) increased accumulation or deposition of extracellular matrix material, (2) decreased protease and stromelysin activities, (3) reorganization of the trabecular meshwork cytoskeleton, (4) increased nuclear size and DNA content, (5) decreased phagocytic activity, and (6) changes in expression of genes that encode specific proteins such as myocilin. Ishibashi, supra. See also Clark A., et al., “Glucocorticoid-induced formation of cross-linked actin networks in cultured human trabecular meshwork cells,” Invest. Ophthalmology & Visual Sci. 35:281-294 (1994); Zhou L., et al., “Glucocorticoids effects on extracellular matrix proteins and integrins in bovine trabecular meshwork cells in relation to glaucoma,” International Journal of Molecular Medicine 1:339-346 (1998); Clark A., et al., “Dexamethasone alters F-actin architecture and promotes cross-linked actin network formation in human trabecular meshwork tissue,” Cell Motility & the Cytoskeleton 60:83-95 (2005); Wordinger R. & Clark A., “Effects of Glucocorticoids on the trabecular meshwork: towards a better understanding of glaucoma,” Progress in Retinal and Eye Research 18:629-667 (1999); Dickerson J., et al., “The effect of dexamethasone on integrin and laminin expression in cultured human trabecular meshwork cells,” Experimental Eye Research 66:731-738 (1998); and Cheng S., et al., “Regulation of avb3 and avb5 integrins by dexamethasone in normal human osteoblastic cells,” Journal of Cellular Biochemistry 77:265-276 (2000), each of which is incorporated herein by reference as if set forth in its entirety.
Interestingly, steroid-induced glaucoma occurs only in a subpopulation of subjects receiving steroid therapy. About 40% of subjects receiving steroids experience an increase in IOP, which in most cases diminishes after therapy ceases. These subjects are called “steroid responders.” Some steroid responders, about 4% to 6%, develop sustained elevated IOP, leading to chronic glaucoma. Steroid-induced glaucoma typically occurs after three to six weeks of steroid use, but in some cases may occur earlier.
In vitro, cultured trabecular meshwork cells or eye organ cultures treated with steroids and cultured trabecular meshwork cells from glaucomatous subjects show a cytoskeletal change in which actin in the trabecular meshwork cells adopts a cross-linked configuration. The cross-linked configuration is referred to as cross-linked actin networks (CLANs) and may make cells more rigid and less responsive to changes in outflow or pressure. CLAN formation is regulated by signaling mechanisms mediated by integrin receptors located on the plasma membrane of mammalian cells. Eye organ cultures are considered by the skilled artisan to be a reliable model system for studying in vivo mammalian eyes and for evaluating therapies for in vivo effectiveness.
Integrins are large glycoproteins having non-covalently linked α- and β-subunits. Integrins are found on virtually all human cells and transmit signals bi-directionally across a cell membrane. As cell surface receptors, integrins participate in a diverse array of biological functions including cellular development, cellular/tissue repair, angiogenesis, inflammation and hemostasis.
Integrin structures and functions are known in the art. See Alpin A., et al., “Signal transduction and signal modulation by cell adhesion receptors: the role of integrins, cadherins, immunoglobulin-cell adhesion molecules, and selectins,” Pharmacological Reviews 50:197-263 (1998); and Brakebusch C. & Fässler R., “The integrin-actin connection, an eternal love affair,” EMBO Journal 22:2324-2333 (2003), each of which is incorporated herein by reference as if set forth in its entirety. At least eighteen isoforms of the α-subunit and eight isoforms of the β-subunit combine to form more than twenty integrin heterodimers having α- and β-subunits. Human trabecular meshwork cells contain the following integrin subunits: α1, α3, α4, α5, α6, αv, β1, β3, β4 and β5. Zhou L., et al., “Expression of receptors in the human trabecular meshwork,” Current Eye Research 19:395-402 (1999). Integrins bind a number of extracellular matrix proteins via an extracellular domain and intact with a variety of tyrosine kinases, adaptor proteins and actin binding proteins via a cytoplasmic tail.
Current treatments for glaucoma include pharmacological and surgical therapies, either alone or in combination. All treatments can have significant side effects. Pharmacological agents, most commonly administered as eye drops, can be used alone or in combination to decrease aqueous humor production or to improve aqueous humor outflow from the eye. β-adrenergic blockers such as timolol, levobunolol and betaxolol decrease aqueous humor production. Side effects of β-adrenergic blockers can include cardiac failure, heart block and bronchospasm. Cholinergic agonists such as pilocarpine, carbachol, and phospholine iodide improve outflow facility from the trabecular meshwork. Side effects of cholinergic agonists can include miosis, brow ache and decreased vision. Carbonic anhydrase inhibitors such as acetazolamide, dorzolamide and brinzolamide decrease aqueous humor production. Side effects of carbonic anhydrase inhibitors can include gastrointestinal upset, malaise, renal stones and aplastic anemia. Non-selective α-agonists such as epinephrine and dipivefrin decrease aqueous humor production and increase trabecular outflow facility. Side effects of a non-selective α-agonists can include pupil dilation, macular edema and tachycardia. Selective α-agonists such as apraclonidine and brimonidine decrease aqueous humor production and increase outflow through the uveoscleral pathway (an alternative, but less utilized, fluid exit pathway to the trabecular meshwork). Side effects of selective α-agonists can include contact allergy and hypotension. Prostaglandin agonists such as latanoprost, travoprost and bimatoprost improve uveoscleral outflow. Side effects of prostaglandins can include iris color change, lash growth and trichiasis. Hyperosmotics such as glycerin (po) and mannitol (iv) establish a concentration gradient that draws excess aqueous humor from the eye. Side effects of using hyperosmotics can include diuresis, cardiovascular overload, renal insufficiency, and stroke, so their use is limited to emergency situations.
When pharmacological agents are unsuccessful in open-angle glaucoma or when a subject presents with closed-angle glaucoma, invasive surgery is indicated. In argon laser trabeculoplasty (ALT), a laser beam is directed at the trabecular meshwork that increases aqueous humor drainage through a mechanism that is not well understood. In laser cyclophotocoagulation, thermal energy applied to the ciliary body destroys the tissue, thereby reducing aqueous humor production. Trabeculectomy establishes a flow route that bypasses the trabecular meshwork so that aqueous humor drains from the anterior chamber just beneath the conjunctiva, the outermost covering of the eye, on the surface of the eye where it is gradually absorbed by blood vessels or diffuses through the conjunctiva. Iridotomy, generally used for closed-angle glaucoma, employs a laser to make an incision in a peripheral area of the iris of the eye to establish a direct aqueous humor flow route between the anterior chamber and the posterior chamber. Iridectomy is similar to iridotomy, but does not employ a laser. In iridotomy, a small section of peripheral iris is surgically excised.
Glaucoma is an increasingly important public health concern, especially in view of the aging of the population. No current treatment satisfactorily and fully addresses onset and control of glaucoma, particularly the particular aspects of steroid-induced glaucoma. In addition, steroids are an effective therapeutic agent in many human diseases, but they can cause unfortunate side effects in a minority of subjects susceptible to steroid-induced glaucoma. Thus, there is a need to develop methods to prevent and treat steroid-induced glaucoma to permit the continued use of steroids as a therapeutic.